We study several plasma membrane proteins that are of medical interest, with a focus on proteins involved in cancer. Lately our interests therein have focused on the role of the organization of proteins in clusters within the membrane.
Tetraspanins are ubiquitous membrane proteins that act as molecular organizers of eukaryotic plasma membranes by assembling specific partner proteins into so-called tetraspanin enriched microdomains (TEMs). We studied the molecular basis for the interaction of tetraspanins with partner proteins using the tetraspanin CD9 and its prototypical partner, the single-span transmembrane protein EWI-F using a compbination of X-ray crustallography of small domains and cryo-EM of liposome embedded complexes.
Six-transmembrane Epithelial Antigen of the Prostate (STEAP) proteins
Six-transmembrane epithelial antigen of the prostate (STEAP) proteins, are a class of metalloreductase enzymes that is upregulated on the cell membranes of several human cancers. STEAPs catalyze the reduction of iron(III) and copper(II), which is a crucial process for the uptake of these metals by mammalian cells. We have solved the 3-D structures of STEAP1 and STEAP4. Our work showed how STEAPs can transfer electrons across the biological membrane and suggest a possible role for STEAP1, which is highly upregulated in several cancers, in modulating the activity of STEAP2-4.
Membrane protein lipidation
Post-translational attachment of palmitate to cysteine residues in membrane proteins is mediated by membrane-embedded palmitoyl-acyl transferases, but the determinants for palmitoylation have remained largely elusive. We used native mass-spectrometry to quantify the palmitoylation states of several membrane proteins. We propose a model in which palmitoylation of cysteines is not dependent on a local sequence motif, but determined by membrane-protein structure, in that it occurs generally on accessible residues within a depth of 8 Å of the lipid bilayer. We showed that lipidation of the tetraspanin CD9 is crucial for its interaction with its partner protein EWI-F and that cysteïne-to-tryptophan mutants are a functional mimic for the lipidated state of CD9, thus providing a a possible biochemical approach to study site-specific membrane-protein lipidation.
Structural basis of R-spondin signalling
R-spondins are potent Wnt-agonists that effect Wnt-driven stem cell proliferation and differentiation. We solved the crystal structures of R-spondin and its complex with the ectodomain of receptor proteins, LGR5, as well as R-spondin in complex with Wnt-antagonist ZNRF3. We show that key binding residues are conserved between R-spondin receptor proteins explaining the promiscuity of R-spondin binding. The data imply the existence of multiple competing complexes that regulate the Wnt-signalling pathway and provide an explanation for several disease causing mutations.